Disease-modifying therapies are needed for the treatment of Rett syndrome (RTT), as there are no current medications that get to the core of disease pathology. Our ongoing work on the design of selective HDAC6 inhibitors (HDAC6i) offers a truly innovative approach to the possible treatment of RTT as it begins to tackle cellular deficits we have identified that arise due to improper MeCP2 function. The preliminary data we have generated demonstrating the ability of an HDAC6i to reduce seizures in Mecp2 mutant mice and improve their rotarod behavioral performance provide a strong foundation for pursuing an HDAC6i-based approach for treating RTT individuals. The innovative aspects of the present undertaking encompass both the chemistry and the biology. Our research program will allow us to refine lead HDAC6i candidates using in vivo and in vitro assays in Mecp2-deficient mice and neuronal cultures, and to obtain valuable ADMET data to aid the translation of this class of molecules to the clinic. Accordingly, our working hypotheses are that upregulated HDAC6 activity contributes to RTT symptomatology, and that pharmacological inhibition of HDAC6 will prevent the development and/or reverse the RTT-like phenotypes in Mecp2-deficient mice. Thus, in our quest for effective RTT therapeutics, we propose a plan of research that encompasses the following aims: Specific Aim 1: Scale-up 10 highly selective Tetrahydroquinoline-based HDAC6i to allow for testing in MeCP2-deficient cells. We have designed and synthesized certain tetrahydroquinoline (THQ)-containing HDAC6is like SW-100 that possess low nM inhibitory activities and in most cases possessing high selectivity (>100-fold) against HDAC6. SW-100 is negative in the Ames test although improvements in its metabolic half-life should be sought. As SW-100 is readily able to penetrate the BBB, and has shown efficacy in Fragile X animals, we believe it is a good candidate for further optimization and study in the Rett animals. Based on in silico ADMET calculations, we have generated 10 new analogs that possess comparable or improved HDAC6i potency. The analogs will be scaled up and screened further for their ability to enhance acetyl-tubulin levels and to improve mitochondrial trafficking in cultured neurons from Mecp2-KO mice. The top 4 compounds showing the best cellular activity will then be submitted for ADMET testing, in order to ensure acceptable brain PK, lack of Ames and hERG activity, low inhibition of the CYP enzymes, etc. See Table X for desired ADMET parameters. Specific Aim 2: Next, after completion of the ADMET tests, the two best compounds from Aim 1 will be studied for their ability to correct the hypo-acetylated tubulin phenotype present in the brains of symptomatic male Mecp2 KO and female Mecp2 heterozygous (Het) mice, and reduce the incidence rates of their Rett-like epileptiform discharge activity. Any signs of adverse events associated would compound toxicity would be noted, although to date the compounds in this class have not raised safety concerns.